title: Analyzing protein-protein interactions by quantitative mass spectrometry.
abstract: Since most cellular processes depend on interactions between proteins, information about protein-protein interactions (PPIs) provide valuable insights into protein function. Over the last years, quantitative affinity purification followed by mass spectrometry (q-AP-MS) has become a powerful approach to investigate PPIs in an unbiased manner. In q-AP-MS the protein of interest is biochemically enriched together with its interaction partners. In parallel, a control experiment is performed to control for non-specific binding. Quantitative mass spectrometry is then employed to compare protein levels in both samples and to exclude non-specific contaminants. Here, we provide two detailed q-AP-MS protocols for pull-downs with immobilized bait proteins or transient transfection of tagged expression constructs. We discuss benefits and limitations of q-AP-MS and highlight critical parameters that need to be considered. The protocols and background information presented here allow the reader to adapt the generic q-AP-MS strategy for a wide range of biological questions.
authors.affiliation: Cell Signaling and Mass Spectrometry Group, Max Delbrück Center for Molecular Medicine, Berlin, Germany.
authors.forename: Florian E
authors.initials: FE
authors.lastname: Paul
authors.forename: Fabian
authors.initials: F
authors.lastname: Hosp
authors.forename: Matthias
authors.initials: M
authors.lastname: Selbach
mesh_terms: D002853:Chromatography, Liquid / Q000379:methods; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans; D007553:Isotope Labeling; D025941:Protein Interaction Mapping / Q000379:methods*; D011506:Proteins / Q000737:chemistry*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D011506:Proteins